Super-resolution microscopy is crucial for imaging sub-wavelength biological structures. However, most techniques rely on nonlinear saturation or stochastic switching of emitters, limiting imaging speed and increasing phototoxicity. Here, we achieve deep super-resolution without employing saturation or stochastic dynamics, instead using a form of spatial mode demultiplexing. By shaping the point-spread function of the emitted light, our Structured Detection Microscope (SDM) redistributes information away from high shot-noise regions of the image, enhancing sensitivity to sub-diffraction emitter separations in two-dimensions and without mode-sorting optics. Implementing SDM within a high-numerical aperture total internal reflection fluorescence microscope, we demonstrate imaging of fluorophores attached to DNA nanorulers with separations as small as 50 nm at resolutions surpassing 40 nm - fivefold below the diffraction limit. This shows that spatial mode demultiplexing can achieve far sub-wavelength resolution and is applicable to biologically relevant samples. By enabling super-resolution biomolecular imaging without emitter saturation and stochasticity, our work opens the door to better understanding biological structure, function and dynamics.
Super-resolution microscopy is crucial for imaging sub-wavelength biological structures. However, most techniques rely on nonlinear saturation or stochastic switching of emitters, limiting imaging speed and increasing phototoxicity. Here, we achieve deep super-resolution without employing saturation or stochastic dynamics, instead using a form of spatial mode demultiplexing. By shaping the point-spread function of the emitted light, our Structured Detection Microscope (SDM) redistributes information away from high shot-noise regions of the image, enhancing sensitivity to sub-diffraction emitter separations in two-dimensions and without mode-sorting optics. Implementing SDM within a high-numerical aperture total internal reflection fluorescence microscope, we demonstrate imaging of fluorophores attached to DNA nanorulers with separations as small as 50 nm at resolutions surpassing 40 nm - fivefold below the diffraction limit. This shows that spatial mode demultiplexing can achieve far sub-wavelength resolution and is applicable to biologically relevant samples. By enabling super-resolution biomolecular imaging without emitter saturation and stochasticity, our work opens the door to better understanding biological structure, function and dynamics.
Recovering both amplitude and phase information from a system is a fundamental goal of optical imaging. At the same time, it is crucial to operate at low photon doses to avoid altering the sample, particularly in biological applications. Quantum imaging provides a powerful route to extract more information per photon than classical techniques, which are ultimately limited by shot-noise. However, the trade-off between quantum noise reduction and spatial resolution has long been regarded as a major obstacle to the application of quantum techniques to small cellular and sub-cellular structures, where they could offer the greatest benefits. Here, we overcome this limitation by demonstrating sub-shot-noise quantitative phase imaging of biological cells based on the transport-of-intensity equation, enabling high-fidelity, label-free imaging of key cellular and sub-cellular features. We achieve high-resolution phase imaging limited only by the numerical aperture, while simultaneously obtaining a resolution-independent quantum advantage. Unlike other quantum imaging approaches, our method operates in a quasi-single-shot, wide-field configuration, retrieves both phase and amplitude information, and does not rely on interferometric measurements, making it intrinsically fast and stable. These results pave the way for the immediate application of sub-shot-noise imaging in biological microscopy.
Recovering both amplitude and phase information from a system is a fundamental goal of optical imaging. At the same time, it is crucial to operate at low photon doses to avoid altering the sample, particularly in biological applications. Quantum imaging provides a powerful route to extract more information per photon than classical techniques, which are ultimately limited by shot-noise. However, the trade-off between quantum noise reduction and spatial resolution has long been regarded as a major obstacle to the application of quantum techniques to small cellular and sub-cellular structures, where they could offer the greatest benefits. Here, we overcome this limitation by demonstrating sub-shot-noise quantitative phase imaging of biological cells based on the transport-of-intensity equation, enabling high-fidelity, label-free imaging of key cellular and sub-cellular features. We achieve high-resolution phase imaging limited only by the numerical aperture, while simultaneously obtaining a resolution-independent quantum advantage. Unlike other quantum imaging approaches, our method operates in a quasi-single-shot, wide-field configuration, retrieves both phase and amplitude information, and does not rely on interferometric measurements, making it intrinsically fast and stable. These results pave the way for the immediate application of sub-shot-noise imaging in biological microscopy.
The resolution of imaging devices is ultimately limited by the diffraction of light. This limit can be circumvented by exploiting near-field phenomena, non-linearities of the sample or specific illumination schemes, which led to the recent explosion of optical super-resolution techniques. However, it would be desirable to reach the nanoscale in the most general way possible, with far-field linear imaging of uniformly illuminated samples. In the last few years, the answer to that came with the field of quantum-inspired super-resolution (QISR), where quantum metrological techniques were used to find novel measurement schemes that, beyond the conventional direct imaging of light, would achieve higher precisions. Interestingly, these ideas would always in some way or another involve transverse spatial modes of light. This thesis presents our work on the subject, with novel theoretical and experimental results. Firstly, we develop the traditional theory of QISR, with has mostly focused on either the estimation of separation of two point sources or on geometric moments of samples. Then, we introduce our own results on optical imaging of general distributions of point sources, in a step to take QISR from toy examples to real imaging scenarios. Secondly, we apply all the concepts studied on a experimental, proof-of-principle implementation of a QISR scheme dubbed Hermite-Gaussian Microscopy (HGM), which uses heterodyne detection with several Hermite-Gaussian modes as local oscillator in order to reconstruct the object sample and achieve super-resolution. We report the techniques necessary for the manipulation of spatial modes of light, as well as our results, where we demonstrated 2D imaging with twofold resolution enhancement beyond the diffraction limit.
Much of our progress in understanding microscale biology has been powered by advances in microscopy. For instance, super-resolution microscopes allow the observation of biological structures at near-atomic-scale resolution, while multi-photon microscopes allow imaging deep into tissue. However, biological structures and dynamics still often remain out of reach of existing microscopes, with further advances in signal-to-noise, resolution and speed needed to access them. In many cases, the performance of microscopes is now limited by quantum effects – such as noise due to the quantisation of light into photons or, for multi-photon microscopes, the low cross-section of multi-photon scattering. These limitations can be overcome by exploiting features of quantum mechanics such as entanglement. Quantum effects can also provide new ways to enhance the performance of microscopes, such as new super-resolution techniques and new techniques to image at difficult to reach wavelengths. This review provides an overview of these various ways in which quantum techniques can improve microscopy, including recent experimental progress. It seeks to provide a realistic picture of what is possible, and what the constraints and opportunities are.
Far-field optical microscopy beyond the Abbe diffraction limit, making use of nonlinear excitation (e.g. STED), or temporal fluctuations in fluorescence (PALM, STORM, SOFI) is already a reality. In contrast, overcoming the diffraction limit using non-classical properties of light is very difficult to achieve due to the fragility of quantum states of light. Here, we experimentally demonstrate superresolution microscopy based on quantum properties of light naturally emitted by fluorophores used as markers in fluorescence microscopy. Our approach is based on photon antibunching, the tendency of fluorophores to emit photons one by one rather than in bursts. Although a distinctively quantum phenomenon, antibunching is readily observed in most common fluorophores even at room temperature. This nonclassical resource can be utilized directly to enhance the imaging resolution, since the non-classical far-field intensity correlations induced by antibunching carry high spatial frequency information on the spatial distribution of emitters. Detecting photon statistics simultaneously in the entire field of view, we were able to detect non-classical correlations of the second and third order, and reconstructed images with resolution significantly beyond the diffraction limit. Alternatively, we demonstrate the utilization of antibunching for augmenting the capabilities of localization-based superresolution imaging in the presence of multiple emitters, using a novel detector comprised of an array of single photon detectors connected to a densely packed fiber bundle. These features allow us to enhance the spatial and temporal resolution with which multiple emitters can be imaged compared with other techniques that rely on CCD cameras.
Intensity correlation microscopy (ICM), which is prominently known through antibunching microscopy or super-resolution optical fluctuation imaging (SOFI), provides super-resolution through a correlation analysis of antibunching of independent quantum emitters or temporal fluctuations of blinking fluorophores. For correlation order m the PSF in the signal is effectively taken to the mth power, and is thus directly shrunk by the factor m. Combined with deconvolution, a close to linear resolution improvement of factor m can be obtained. Yet, analysis of high correlation orders is challenging, which limits the achievable resolutions. Here we propose to use three dimensional structured illumination along with mth-order correlation analysis to obtain an enhanced scaling of up to m + m = 2m. Including the stokes shift or plasmonic sub-wavelength illumination enhancements beyond 2m can be achieved. Hence, resolutions far below the diffraction limit in full 3D imaging and with already low correlation orders, can potentially be achieved. Since ICM operates in the linear regime our approach may be particularly promising for enhancing the resolution in biological imaging at low illumination levels.